PROJECT SUMMARY Although Staphylococcus aureus is a major human pathogen that causes an enormous burden of disease, we have very few strategies for preventing infection or controlling its spread. In the 1960's it was a widespread practice to inoculate neonates with a relatively benign strain of S. aureus called 502A as a strategy to combat outbreaks of more virulent staphylococcal strains, a practice referred to as bacterial interference. Thousands of neonates were exposed to 502A with very little associated morbidity and mortality, but despite its relatively good safety record, the practice was abandoned in the 1970's due to safety concerns. The basic mechanisms of bacterial interference were never investigated. We recently showed that 502A elicits a very different cytokine response compared to other epidemic and virulent strains. Although it is less invasive, host cells exposed to 502A have increased levels of type I Interferons as well as several other proinflammatory cytokines (eg., TNFa, IL-6) compared to exposure to other S. aureus strains. Our hypothesis is that S. aureus strains that stimulate 502A-like immune responses protect from colonization and subsequent infection by inducing the immune system to reject other strains. Understanding the mechanism of 502A bacterial interference could allow us to design strategies for protection against colonization and infection with virulent or antibiotic resistant strains. It may also lead to strategies for decolonization in the setting of recurrent S. aureus infection or in intensive care units where universal decolonization using antimicrobials has been show to decrease mortality. We combine a clinical approach with basic molecular research, aiming to assess the feasibility of a bacterial interference strategy in the clinical context. The study team integrates expertise in clinical S. aureus epidemiology, host innate immunology, and genomics and microbiome studies. Aim 1 uses the natural diversity in S. aureus strains found in human populations to prospectively assess the correlation of colonization with 502A-like strains and possible protection. Aim 2 uses model systems to assess the response of the immune system to 502A-like strains as well as their protective effect. Aim 3 examines the possibility of using a virulence-attenuated strain for bacterial interference and decolonization.